Extended Data Fig. 1: The first two approaches to construct linear plasmids based on lytic phage and temperate phage in B. subtilis.

(a) Linear plasmid system construction using lytic phage φ29 replication system. During the phage φ29 lytic cycle, phage φ29 first injects its linear double-stranded DNA (dsDNA) genome into the cell. Then, it replicates the phage genome and synthesizes phage capsid and tail proteins. In addition, cells are lysed, and progeny phages are released after packaging into progeny phage particles. Developing linear plasmids in B. subtilis was tested by expressing linear dsDNA replication machinery proteins and rationally designing linear plasmids. TP, terminal proteins. DNAP, DNA polymerase. (b) Transferring the dormant state phage to B. subtilis by protoplast fusion of B. thuringiensis-harboring temperate phage GIL01 and B. subtilis. Linear plasmid system construction using temperate phage GIL01 replication system. B. thuringiensis temperate phage GIL01/GIL16 are capable of persistent intracellular dormancy unless DNA damage occurs. Therefore, it potentially can be used as a linear plasmid in B. subtilis.